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Selection of alternative 5' splice sites: role of U1 snRNP and models for the antagonistic effects of SF2/ASF and hnRNP A1.
Mol Cell Biol. 2000 Nov; 20(22):8303-18.MC

Abstract

The first component known to recognize and discriminate among potential 5' splice sites (5'SSs) in pre-mRNA is the U1 snRNP. However, the relative levels of U1 snRNP binding to alternative 5'SSs do not necessarily determine the splicing outcome. Strikingly, SF2/ASF, one of the essential SR protein-splicing factors, causes a dose-dependent shift in splicing to a downstream (intron-proximal) site, and yet it increases U1 snRNP binding at upstream and downstream sites simultaneously. We show here that hnRNP A1, which shifts splicing towards an upstream 5'SS, causes reduced U1 snRNP binding at both sites. Nonetheless, the importance of U1 snRNP binding is shown by proportionality between the level of U1 snRNP binding to the downstream site and its use in splicing. With purified components, hnRNP A1 reduces U1 snRNP binding to 5'SSs by binding cooperatively and indiscriminately to the pre-mRNA. Mutations in hnRNP A1 and SF2/ASF show that the opposite effects of the proteins on 5'SS choice are correlated with their effects on U1 snRNP binding. Cross-linking experiments show that SF2/ASF and hnRNP A1 compete to bind pre-mRNA, and we conclude that this competition is the basis of their functional antagonism; SF2/ASF enhances U1 snRNP binding at all 5'SSs, the rise in simultaneous occupancy causing a shift in splicing towards the downstream site, whereas hnRNP A1 interferes with U1 snRNP binding such that 5'SS occupancy is lower and the affinities of U1 snRNP for the individual sites determine the site of splicing.

Authors+Show Affiliations

Department of Biochemistry, University of Leicester, Leicester LE1 7RH, United Kingdom. eci@le.ac.ukNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

11046128

Citation

Eperon, I C., et al. "Selection of Alternative 5' Splice Sites: Role of U1 snRNP and Models for the Antagonistic Effects of SF2/ASF and hnRNP A1." Molecular and Cellular Biology, vol. 20, no. 22, 2000, pp. 8303-18.
Eperon IC, Makarova OV, Mayeda A, et al. Selection of alternative 5' splice sites: role of U1 snRNP and models for the antagonistic effects of SF2/ASF and hnRNP A1. Mol Cell Biol. 2000;20(22):8303-18.
Eperon, I. C., Makarova, O. V., Mayeda, A., Munroe, S. H., Cáceres, J. F., Hayward, D. G., & Krainer, A. R. (2000). Selection of alternative 5' splice sites: role of U1 snRNP and models for the antagonistic effects of SF2/ASF and hnRNP A1. Molecular and Cellular Biology, 20(22), 8303-18.
Eperon IC, et al. Selection of Alternative 5' Splice Sites: Role of U1 snRNP and Models for the Antagonistic Effects of SF2/ASF and hnRNP A1. Mol Cell Biol. 2000;20(22):8303-18. PubMed PMID: 11046128.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Selection of alternative 5' splice sites: role of U1 snRNP and models for the antagonistic effects of SF2/ASF and hnRNP A1. AU - Eperon,I C, AU - Makarova,O V, AU - Mayeda,A, AU - Munroe,S H, AU - Cáceres,J F, AU - Hayward,D G, AU - Krainer,A R, PY - 2000/10/25/pubmed PY - 2001/2/28/medline PY - 2000/10/25/entrez SP - 8303 EP - 18 JF - Molecular and cellular biology JO - Mol Cell Biol VL - 20 IS - 22 N2 - The first component known to recognize and discriminate among potential 5' splice sites (5'SSs) in pre-mRNA is the U1 snRNP. However, the relative levels of U1 snRNP binding to alternative 5'SSs do not necessarily determine the splicing outcome. Strikingly, SF2/ASF, one of the essential SR protein-splicing factors, causes a dose-dependent shift in splicing to a downstream (intron-proximal) site, and yet it increases U1 snRNP binding at upstream and downstream sites simultaneously. We show here that hnRNP A1, which shifts splicing towards an upstream 5'SS, causes reduced U1 snRNP binding at both sites. Nonetheless, the importance of U1 snRNP binding is shown by proportionality between the level of U1 snRNP binding to the downstream site and its use in splicing. With purified components, hnRNP A1 reduces U1 snRNP binding to 5'SSs by binding cooperatively and indiscriminately to the pre-mRNA. Mutations in hnRNP A1 and SF2/ASF show that the opposite effects of the proteins on 5'SS choice are correlated with their effects on U1 snRNP binding. Cross-linking experiments show that SF2/ASF and hnRNP A1 compete to bind pre-mRNA, and we conclude that this competition is the basis of their functional antagonism; SF2/ASF enhances U1 snRNP binding at all 5'SSs, the rise in simultaneous occupancy causing a shift in splicing towards the downstream site, whereas hnRNP A1 interferes with U1 snRNP binding such that 5'SS occupancy is lower and the affinities of U1 snRNP for the individual sites determine the site of splicing. SN - 0270-7306 UR - https://www.unboundmedicine.com/medline/citation/11046128/Selection_of_alternative_5'_splice_sites:_role_of_U1_snRNP_and_models_for_the_antagonistic_effects_of_SF2/ASF_and_hnRNP_A1_ L2 - http://mcb.asm.org/cgi/pmidlookup?view=long&pmid=11046128 DB - PRIME DP - Unbound Medicine ER -